Abstract Agenesis of the corpus callosum (CC), the large white matter tract connecting the cerebral hemispheres, occurs in about 1:4000 individuals and in 3-5% of patients with a developmental brain disorder, in particular intellectual disability or autism spectrum disorder (ASD). CC agenesis is also present in approximately 10% of individuals with CHARGE syndrome, a multiple anomaly developmental disorder caused by heterozygous mutations in CHD7, which encodes a chromodomain helicase protein. Importantly, because defects in CC development can only be ascertained by post-mortem analysis or structural MRI, which is not medically indicated in CHARGE syndrome, they may be significantly under-diagnosed. The contribution of CC agenesis to the intellectual deficits and autistic behaviors characteristic of CHARGE syndrome remains unexplored. In preliminary studies, I identified a novel, highly-penetrant CC agenesis phenotype in a model of CHARGE syndrome, wherein presumptive callosal axons failed to cross the midline, formed aberrant Probst bundles and were misrouted to the septum. Subsequent conditional deletion of Chd7 suggested that CC agenesis developed non-cell autonomously, likely involving Chd7 haploinsufficiency in a novel cell population via an unexplored mechanism. In additional fate-mapping studies, I identified the choroid plexus and the pia mater as two potential tissues that can contribute to CC agenesis in the CHARGE model. Interestingly, emerging work has suggested the meninges, specifically the pia, to be a critical controller of midline and callosal development through signaling and physical contact with neurons. However, the contribution of pial development to the CC agenesis and additional axon tract defects in CHARGE syndrome has never been explored. My central hypothesis is that agenesis of the corpus callosum in CHARGE syndrome is the result of non-cell autonomous deficits in the development or function of the midline pia mater. In this proposal, I will test this hypothesis by (1) systematic analysis of the forebrain midline populations necessary for callosum development in the animal model of CHARGE syndrome. Leveraging available mouse lines, I will (2) generate additional conditional deletions of Chd7 to dissect the potential contribution of meningeal and choroid plexus tissue to the CC phenotype. And (3) I will analyze changes in the transcriptomes of pia mater and choroid plexus cells during callosal development to identify downstream genes that may underpin this phenotype. Together, this work is expected to lead to a mechanistic understanding of an underexplored phenotype that may contribute to key aspects of the CHARGE syndrome phenotype. This line of scientific pursuit is perfectly aligned with my ultimate career goal to establish an independent research program to seek a functional and translational understanding of the genetic mechanisms underlying developmental brain disorders. As detailed in the training plan, the proposed work will integrate scientific, technical, and clinical training, as well as career development opportunities, that will propel me towards a career as a clinician-scientist with active research.